Cloning And Expression Of Novel Epoxide Hydrolases And Their Performance In Biocatalytic Resolution Of Phenyl Glycidyl Ethers

With the development of genome sequencing, more and more whole genomes are released in the public database, which facilitates the cloning of target genes. The predicted epoxide hydrolase (EH) genes in Populus trichocarpa and Bacillus megaterium were successfully cloned in the study. The investigation of substrate scope indicated that, BMEH could preferentially hydrolyze aryl glycidyl ethers, yielding the optically active epoxides or vicinal diols. For phenyl glycidyl ether as a model substrate, BMEH exhibited high (R)-selectivity (E =58), which is the highest among all the known native EHs to date. Generally speaking, the highest enantioselectivities (E> 200) were achieved in the bioresolution of ortho-substituted phenyl glycidyl ethers and para-nitrostyrene oxide. Noteworthily, a reversed (S)-enantiopreference was observed unexpectedly for the ortho-nitro-phenyl glycidyl ether (oNPGE). More interestingly, excellent E values (>200) were observed in both the cases of ortho-and meta-nitro-substituted phenyl glycidyl ethers, although the enantiopreference was opposite. Subsequent study of regioselectivity showed that BMEH attacks very preferentially at the less substituted carbon atom. However, it was noticed with a surprise that the oxirane ring was preferentially attacked at the more substituted carbon atom for oNPGE. Molecular docking was then performed to uncover the orign of reverted enantioselectivity. As a proof-of-concept, five enantiopure epoxides (>99%ee) were obtained in high yields, and a gram-scale preparation of (S)-ortho-methylphenyl glycidyl ether was then successfully performed within a few hours, indicating that BMEH is an attractive biocatalyst for the efficient preparation of optically active epoxides and vicinal diols.